Liver Mitochondrial Permeability Transition Research Articles

L-Deprenyl as an inhibitor of menadione-induced permeability transition in liver mitochondria

l-Deprenyl, an inhibitor of mitochondrial monoamine oxidase B (MAO B), inhibits the swelling of liver mitochondria induced by the pro-oxidant 2-methyl-1,4-naphtoquinone with a K i dependent on quinone concentration. l-Deprenyl also inhibits the collapse of membrane potential, cation efflux, pyridine nucleotide oxidation and cytochrome c release, all events which accompany the osmotic change and are typical of membrane permeability transition induction, thus emphasizing the inhibitory effect of the drug on this phenomenon. Results show that this inhibition is not due to the effect of l-deprenyl on monoamine oxidase activity but is most likely due to a direct interaction of the drug with the pore forming structures. It is here proposed that l-deprenyl, being a propargylamine, at physiological pH has a protonated amino group able to interact with critical aromatic or anionic amino acidic residues. As a consequence, the opening of the transition pore is prevented. These results indicate a more generalized protective effect of l-deprenyl on mitochondrial functions, involving the inhibition of membrane permeability transition induced not only by the oxidation of substrates of MAO B, but also by pro-oxidant agents such as 2-methyl-1,4-naphtoquinone, which does not involve MAO B activity.

Limitations of Cyclosporin A Inhibition of the Permeability Transition in CNS Mitochondria

Activation of the mitochondrial permeability transition may contribute to excitotoxic neuronal death (Ankarcrona et al., 1996; Dubinsky and Levi, 1998). However, cyclosporin A (CsA), a potent inhibitor of the permeability transition in liver mitochondria, only protects against neuronal injury by limited doses of glutamate and selected ischemic paradigms. The lack of consistent CsA inhibition of the mitochondrial permeability transition was analyzed with the use of isolated brain mitochondria. Changes in the permeability of the inner mitochondrial membrane were evaluated by monitoring mitochondrial membrane potential (Deltapsi), using the distribution of tetraphenylphosphonium, and by monitoring mitochondrial swelling, using light absorbance measurements. Metabolic impairments, large Ca(2+) loads, omission of external Mg(2+), or low doses of palmitic acid or the protonophore FCCP exacerbated Ca(2+)-induced sustained depolarizations and swelling and eliminated CsA inhibition. BSA restored CsA inhibition in mitochondria challenged with 50 microm Ca(2+), but not with 100 microm Ca(2+). CsA failed to prevent Ca(2+)-induced depolarization or to repolarize mitochondria when mitochondria were depolarized excessively. Similarly, CsA failed to prevent mitochondrial swelling or PEG-induced shrinkage after swelling when the Ca(2+) challenge produced a strong, sustained depolarization. Thus in brain mitochondria CsA may be effective only as an inhibitor of the permeability transition and the Ca(2+)-activated low permeability state under conditions of partial depolarization. In contrast, ADP plus oligomycin inhibited both permeabilities under all of the conditions that were tested. In situ, the neuroprotective action of CsA may be limited to glutamate challenges sufficiently toxic to induce the permeability transition but not so severe that mitochondrial depolarization exceeds threshold.

Cholestasis confers resistance to the rat liver mitochondrial permeability transition

Background & Aims: Bile salts can cause hepatocyte death by inducing the mitochondrial permeability transition (MPT). However, the slow progression of human cholestatic liver diseases suggests that hepatocytes adapt to resist the MPT. Bcl-x, a protein, and increased mitochondrial cardiolipin, a membrane lipid, elevate the threshold for the MPT. Our aims were to determine if liver mitochondria become resistant to the MPT during cholestasis and, if so, if the resistance is mediated by Bcl-x and/or increased cardiolipin. Methods: Hepatocytes and liver mitochondria were obtained from bile duct–ligated (BDL) rats and sham-operated rats (control). Results: After addition of glycochenodeoxycholate (GCDC), the magnitude of the MPT was reduced in mitochondria from BDL rats vs. controls. Although Bcl-x L was not increased, mitochondrial cardiolipin content was significantly greater in BDL rats vs. controls. Cell viability was also increased in hepatocytes from BDL rats vs. controls after treatment with GCDC. Feeding BDL rats a fatty acid–deficient diet prevented the increase in mitochondrial cardiolipin content; mitochondria and hepatocytes from these rats were susceptible to the MPT and hepatocellular death by GCDC. Conclusions: These data suggest that an increase in mitochondria cardiolipin content occurs during cholestasis as an adaptive phenomenon to resist cell death by the MPT. GASTROENTEROLOGY 1998;115:693-701

The Contribution of Endogenous Polyamines to the Permeability Transition of Rat Liver Mitochondria

Spermine and spermidine are present in rat liver mitochondria at the concentrations of 3.5 and 1.8 nanomoles/mg protein, respectively. Addition of 50 μM Ca2+and 1 mM Pi to respiring mitochondria induces, concomitantly with mitochondrial swelling and Mg2+efflux, a consistent release of polyamines, which is either prevented by cyclosporin A or Mg2+. Addition of 0.1 mM spermine to mitochondria deenergized by Ca2+and phosphate restores, like 1 mM Mg2+, transmembrane potential at the physiological level, while either cyclosporin A or ADP are ineffective. The presence of polyamines in the matrix space should be considered relevant, like Mg2+, in controlling the permeability transition of liver mitochondria.

Effects of the anaesthetic propofol on the calcium-induced permeability transition of rat heart mitochondria: direct pore inhibition and shift of the gating potential

Mitochondrial calcium exchanges are involved in intracellular calcium homeostasis and in the contraction-relaxation process in myocytes. The calcium-induced permeability transition of the heart mitochondria inner membrane appears to be an important calcium efflux mechanism involved in some physiological and pathological situations. The negative inotropic effect of the anaesthetic propofol results in part from a decrease in intracellular calcium availability. Thus, this study evaluates the effects of propofol on calcium transport and permeability transition of heart mitochondria. The propofol-inhibition of the permeability transition of liver mitochondria was previously investigated [Eriksson, O. (1991) FEBS Lett. 279, 45–48] in such conditions that its uncoupling effect was not taken into account. We show here that propofol uncoupling results in a decrease in calcium uptake rate which could in part explain the decreased permeability transition rate. However, comparison of equipotent uncoupling concentrations of propofol and carbonylcyanide m-chlorophenylhydrazone reveals that beyond this uncoupling effect, propofol has a direct inhibitory action on the permeability transition pore, concomittant with a shift of its gating potential.

Inhibition of Rat Liver Mitochondrial Permeability Transition by Respiratory Substrates

The mitochondrial inner membrane can undergo a permeability increase known as "permeability transition" elicited by Ca2+ and several other inducing agents. In general, the condition of oxidative stress acts as an inducer, at variance with antioxidants and reducing agents that inhibit the permeability transition. The action of mitochondrial respiratory substrates in preventing the permeability transition induced by Ca2+ and phosphate was examined; pyruvate, isocitrate, and glutamate proved to be particularly effective. The effect of substrates was evident also in the presence of an uncoupler, and, in addition, they were able to counteract the swelling stimulated by acetoacetate and tert-butylhydroperoxide. In the presence of various pyridine nucleotide-dependent substrates, mitochondria are able to reduce the disulfide 5,5′-dithiobis(2-nitrobenzoic acid) (DTNB) to an extent far larger than that calculated from the theoretical amount of total mitochondrial thiol groups, indicating the occurrence of a catalytic system. Similarly, the enzymes of the mitochondrial matrix in the presence of either NADH or NADPH are able to reduce DTNB. The results are discussed considering the existence of a close redox communication between pyridine nucleotides and membrane thiol groups, possibly mediated by dithiols such as thioredoxin and lipoic acid.

Spermine Inhibition of the Permeability Transition of Isolated Rat Liver Mitochondria: An Investigation of Mechanism

Mitochondria that have accumulated Ca 2+ can be induced to undergo a permeability transition: the inner membrane becomes nonselectively permeable to small (<1500 Da) solutes. The molecular mechanism(s) underlying this transition, which is Ca 2+-dependent and cyclosporin A-sensitive, has yet to be clearly elucidated. Our laboratory has recently identified the polyamine spermine as an inhibitor of the permeability transition of isolated rat heart mitochondria. In this study, we have used spermine, in combination with a series of triggering agents, to clarify several mechanistic details of the transition process in isolated rat liver mitochondria. Mitochondrial swelling was monitored as an indicator of transition occurrence. Our results indicate that: (1) spermine inhibits the permeability transition of isolated rat liver mitochondria; (2) the sensitivity of the permeability transition of liver mitochondria to spermine is highly dependent on the ionic composition of the assay medium; (3) K + interacts with a site outside the mitochondria to decrease spermine effectiveness; (4) spermine likewise acts at an external site; and (5) the Ca 2+ uniporter in its inactive form is not the protein responsible for mediating the permeability transition.

Cyclosporin A is a potent inhibitor of the inner membrane permeability transition in liver mitochondria

The immunosuppressive peptide cyclosporin A is a powerful inhibitor of the Ca2+-dependent permeability transition in rat liver mitochondria. When swelling is used to monitor the transition, the inhibitor is effective regardless of whether N-ethylmaleimide, Hg2+, WY-14643, t-butyl hydroperoxide, oxalacetate, rhein, phosphate, phosphoenolpyruvate, or ruthenium red plus uncoupler is used as the inducing agent. Twenty-five to fifty pmol/mg protein of cyclosporin A reduces the swelling response by 50% with complete inhibition obtained at about 150 pmol/mg protein. The compound, which does not inhibit Ca2+ uptake or mitochondrial phospholipase A2, is effective when added before or after the transition promoting agent. These findings, together with the shape of the inhibition dose-response curve, suggest that cyclosporin A essentially titrates a mitochondrial component which is present at 80-90 pmol/mg protein. It is proposed that this component is a solute unselective, regulated pore or a factor involved in controlling such a structure.